Breast cancer progression is driven by deregulated proliferation, the acquisition of inappropriate invasiveness and mitotic defects, in addition to other phenotypic changes. One molecule known to cause progression of breast cancer cells to more invasive phenotypes is p21-activated kinase 1 (Pak1), a nodular kinase activated by a variety of cell surface or intracellular signals. Despite the remarkable growth of new information on the biology of Pak1 in the last decade, we still do not know how Pak1 participates in essential, regulatory events which ensure S-phase progression and controlled mitosis and how it could significantly influence the status of polyploidy in breast cancer The purpose of this project is to clarify and define for the first time the significant role of signal-dependent stimulation of Pak1 in the G1-to-S transition and in mitotic progression, in both physiological and aberrant cell proliferation. These studies are designed to offer a molecular explanation for the well-documented effect of Pak1 dysregulation at two critical points in cell cycle progression. In addition, we will test the hypothesis that E2F1 and HSF1 transcriptional factors are determinants of Pak1 functions at the G1/S phase transition and during mitosis using functionally relevant, genetic mouse models. And finally, we will evaluate and the prognostic significance of these emerging molecules in breast cancer progression. The rationale for this proposal is drawn from a number of findings from the PI's laboratory supported by R01-CA090970-09, for which we are seeking renewal. These findings to suggest that Pak1 signaling plays an inherent role in controlling the G1-to-S phase transition and mitotic progression via E2F1 or HSF1 respectively, and that E2F1 and HSF1 might be novel determinants of Pak1 functions in breast cancer cells. Our testable hypotheses are: Deregulation of Pak1 signaling stimulates E2F1- and HSF1-dependent pathways; and consequently, deregulation of Pak1 signaling confers deregulated G1-to-S-transition and mitotic progression in breast cancer cells. To address these hypotheses, our Specific Aims are to: To address these hypotheses, our Specific Aims are to: (1) Investigate the mechanism by which Pak1 regulates the G1-to-S progression by defining the impact of Pak1-phosphorylation upon the functions of E2Fs; (2) Determine the role of HSF1 in the nuclear accumulation of activated Pak1 and in mitotic progression of breast cancer cells; and (3) To study the role of E2F1 or HSF1 as determinant of Pak1 function in the G1/S transition and mitotic progression in physiological relevant model systems. Our proposed research is significant as the knowledge gained here may reveal Pak1 signaling-dependent regulation of the E2F family and HSF1 transcriptional factors for the first time, and thus, defining novel Pak1 functions in the G1-to-S-transition and mitotic progression of breast cancer cells. Additionally, this work is innovative because we will start understanding the principles by which Pak1 regulates transcription of a subset of genes with roles in the S-phase and mitosis. PUBLIC HEALTH RELEVANCE: Despite the remarkable growth of new information on the biology of p21-activated kinase 1 (Pak1) in breast cancer cells, we still do not know how Pak1 participates in essential, regulatory events which ensure S-phase progression and controlled mitosis and how it could significantly influence the status of polyploidy in breast cancer. The purpose of this project is to clarify and define for the first time the significant role of signal-dependent stimulation of Pak1 in the G1-to-S transition and in mitotic progression, in both physiological and aberrant cell proliferation.